The polymorphic lipophoshoglycan (LPG) present on the surface of Leishmania promastigotes mediates binding to the sand fly vector midgut, and accounts for species restricted vector competence. The LPGs of L. major strains from West Africa were found to lack side chains that terminate in beta-linked galactose, explaining their failure to bind to midguts of P. papatasi, and their reliance on P. duboscqi for natural transmission. An L. major strain from West Africa was transfected with a family of L. major genes that encode galactosyl transferases. Transgenic parasites expressing one or two beta 1-3 Gal side chain residues were rescued for survival in P. papatasi, though transfectants expressing long chain poly gals were not. Screening of L. major LPG structures from an extensive number of isolates from Africa, the Middle East, Afghanistan, South Asia and Southern Russia revealed a remarkable elongation in side chain length moving from West to East in the L. major endemic zones, suggesting adaption to midgut receptor polymorphisms expressed within the P. papatasi species complex. [unreadable] A major effort has been directed toward quantification of the natural transmission dose of Leishmania to mammalian host during the bite of an infected sand fly, which is currently unknown for any natural sand fly vector. The quantitative data are needed to define the dose of parasites that should be used when designing experimental needle infections intended to mimic natural transmission, and revealing natural dose ranges that may influence clinical outcome. A real-time PCR-based method was developed to determine the number of Leishmania promastigotes transmitted to a live host during blood feeding by a single infected sand flies. As expected, flies bearing heavier infections tended to transmit a larger number of parasites while feeding. Well-infected flies never fed to completion and frequently took a blood-meal that less than half filled the mid-gut. Transmission to mice could be detected from flies that had taken no detectable blood-meal at all. Dose ranges so far documented are 1000-5000 parasites, though the sensitivity of parasite detection in host tissue below 1000 needs to be improved. [unreadable] Transmission by sand fly bite is being used for the first time to evaluate the efficacy of experimental vaccines against leishmaniasis. C57BL/6 mice were vaccinated with live parasites or a highly effective killed vaccine, ALM+CpG, and exposed to infected sand fly bites or needle challenge. In live-vaccinated mice, protective antigen-specific IFN-gamma-producing CD4+ T cells were found at the site of sand fly bite within 24 hours and a substantial increase was observed by 72 hours. In contrast, recruitment of IFN-gamma-producing CD4+ T cells to the site of sand fly bite in ALM+CpG vaccinated mice was delayed until 1 week following challenge, and their numbers were dramatically reduced. Four weeks post-challenge, ALM+CpG vaccination showed strong protection against needle inoculation, but no protection against infected sand fly bite, whereas live-vaccination protected against both. Sand fly challenge was associated with early recruitment of neutrophils and macrophages to the site of bite that may have compromised the expression of immunity in the ALM+CpG immunized mice. By contrast, the rapid recruitment of large numbers of effector cells to the site of sand fly bite in the live vaccinated mice appeared to overcome these suppressive conditions. These observations are highly relevant to vaccine design and development as they identify critical correlates for vaccine efficacy against L. major.